Electrophoretic matrix staining composition and method

ABSTRACT

Staining compositions and methods for use with a matrix, such as an electrophoretic gel, containing separated biopolymers. The compositions including an acid, an organic solvent, and a generally planar, fluid-permeable gel contact sheet consisting primarily of a non-cellulosic material. The acid and organic solvent may be sorbed to the fluid-permeable gel contact sheet, or may be sorbed to a layers contactable with a non-gel contact side of the gel contact sheet. In one embodiment, the composition is a source electrophoretic stain composition including a staining reagent. In one embodiment, the composition is a sink electrophoretic stain composition including a sorbent.

This application claims priority to U.S. Patent Application Ser. No.61/882,951 filed Sep. 26, 2013 which is expressly incorporated byreference in its entirety.

Compositions, also referred to as consumables, and method for stainingbiopolymers such as proteins, nucleic acids, oligosaccharides, etc. in amatrix using an electric field. The staining composition hasfluid-permeable sheet(s) that is/are brought into contact with thematrix. In one embodiment the fluid-permeable sheet(s) is/are primarilya non-cellulosic material such as polyester or polyester blend, and inone embodiment is/are a non-polymeric non-cellulosic material, e.g.,natural fiber materials. The composition contains a source contact sheetincluding a staining reagent and a sink contact sheet, where thestaining reagent interacts, associates, and/or binds with a biopolymer.The inventive method stains biopolymers using an electric fieldemploying these source and sink contact sheets. In embodiments, thecompositions are highly absorbent, liquid retaining, and fluidpermeable, which maximize contact with the matrix by providing at leastone of a smooth surface, uniform consistency, and pliability forimproved conformation with the matrix surface. Without being held to asingle theory, maximization of the contact between the disclosedcompositions and the matrix provides uniform transfer of current by theelectric field and results in improved staining and destaining.

U.S. Patent Application Publication No. 2010/0326828 discloses acomposition and method for staining biopolymers after gelelectrophoresis that uses paper materials, specifically blotting paper.A staining solution, including a stain such as Coomassie Blue R-250 orG-250, is either pre-absorbed or applied to a porous material such asthe blotting paper. The solution-containing blotting paper is positionedto contact one side of an electrophoresis gel, and a second blottingpaper, lacking the stain, is positioned to contact the other side of thegel forming an assembly. This assembly is arranged between a pair ofelectrodes and a DC voltage is applied to the electrodes and across theassembly. The voltage difference between the electrodes causes the stainto migrate out of the solution-containing blotting paper and into thegel, where the stain binds to biopolymers in the gel matrix. The voltagedifference is maintained to cause any unbound stain to migratecompletely through the gel and into the second blotting paper,destaining the gel matrix but not destaining the biopolymers in thematrix. The composition and method replace passive, diffusion-drivenprocesses for staining electrophoretic separations; these may requireseveral hours to complete, while the active electrically-driven stainingprocess requires as short a time as six minutes.

As stated, the '828 application uses paper materials such as filterpaper, blotting paper, paper pads, etc. that are cellulosic products.The commercially available eStain® 2.0 Protein Staining System(GenScript USA, Inc., Piscataway N.J.) is based on a cotton cellulosefiber paper.

The use of these cellulosic materials in the electrically-drivenstaining method yields exhibit undesirable effects, such as highbackgrounds and/or blotchy backgrounds in electrophoresis gels. Withoutbeing limited to a specific theory, such undesirable backgrounds resultfrom the non-uniform and relatively rigid surfaces of the papermaterials that do not provide a substantially maximal and/or uniformcontact with the matrix surface. Although the method disclosed in the'828 application has increased speed, its resultant high backgroundforces a trade-off between staining time and low background levels,similar to diffusion-driven processes, which are often performedovernight to essentially completely remove background and improve theprecision and/or accuracy of automated fingerprint detection methods andquantitation methods.

The disclosed compositions and methods use the reduced staining timeachieved with electrically-driven staining method, as well as alsoproviding substantially improved stain backgrounds that are both low anduniform. The disclosed compositions and methods are useful for and areused in, e.g., semi-automated or fully-automated electrophoreticseparation processes used in electrophoretic fingerprint detectionmethods and quantitation methods.

One embodiment is an electrophoretic stain composition including asource sheet, which includes embodiments where the source sheet is aplurality of sheets, containing a stain solution, i.e., a staindissolved in a suitable solvent, and a sink sheet or sheets containing asuitable solution, such as a destain solution. The sheet or sheets aregenerally planar, fluid-permeable contact sheet primarily of anon-cellulosic material or a cellulosic blend. The stain solution and/ordestain solution may be sorbed to the fluid-permeable contact sheet, ormay be sorbed to a reservoir layer contactable with a non-matrix contactside of the contact sheet. In one construction, the source sheet orsheets is a source electrophoretic stain composition including a stain.In another construction, the sink sheet or sheets is a sinkelectrophoretic stain composition including a destain solution.

One embodiment is an electrophoretic stain kit. The kit includes a firstcomposition of an acid, an organic solvent, a staining reagent, i.e., astain, and a first generally planar, fluid-permeable contact sheetprimarily of a non-cellulosic material. Any one or more of the acid,organic solvent, and stain may be sorbed to the fluid-permeable contactsheet, or may be sorbed to a reservoir layer contacting a non-matrixcontact side of the first contact sheet. The kit also includes a secondcomposition of a suitable solution, such as a destain solution, and asecond generally planar, fluid-permeable contact sheet primarily of anon-cellulosic material. A sorbent may be a non-primary component of thesecond gel contact sheet, or may be a sorbent layer contacting anon-matrix contact side of the second contact sheet.

One embodiment is a method of staining a matrix such as anelectrophoretic gel containing a biopolymer. The method includes (a)preparing or constructing an assembly of (i) an electrophoretic gelcontaining the biopolymer (ii) a first generally planar, fluid-permeablecontact sheet primarily of a non-cellulosic material having a matrixcontact side in direct contact with a side of the electrophoretic geland (iii) a second generally planar, fluid-permeable contact sheetprimarily of a non-cellulosic material having a matrix contact side indirect contact with an opposite side of the electrophoretic gel; (b)positioning the assembly between a first electrode and a secondelectrode and in electrical contact with the electrodes; and (c)applying a DC current to the electrodes and across the assembly. Beforeapplying the current, the first contact sheet is provided with astaining solution, and the second contact sheet is provided with atleast a suitable solution, such as a destain solution.

In each embodiment, the generally planar, fluid-permeable contact sheetsconsist primarily of a non-cellulosic material, but are not necessarilycellulose-free. For example, the gel contact sheets may be manufacturedfrom polyester or a polyester cellulose blend, with the sheet beingprincipally polyester but including a lesser fraction of cellulose,e.g., non-woven cellulose fiber.

BRIEF DESCRIPTION OF THE DRAWINGS

The patent or application file contains at least one drawing executed incolor. Copies of this patent or patent application publication withcolor drawing(s) will be provided by the Office upon request and paymentof the necessary fee.

FIGS. 1A, B, C, D are pictures of electrophoresis gels processed witheStain® 2.0 Protein Staining System using the supplied blotting paper asa gel-contacting stain solution carrier and destaining sink contactsheet (FIGS. 1A, 1B), and gels stained using a conventional Coomassiestaining protocol (FIGS. 1C, 1D).

FIGS. 2A-B compare electrical resistance (FIG. 2A) and temperaturegenerated (FIG. 2B) during the electrophoretic staining procedure usingfilter paper or polyester cellulose material as a gel-contacting stainsolution carrier and destain sink contact sheet.

FIGS. 3A-B are pictures of electrophoresis gels processed using theprocedure of U.S. Patent Application Publication No. 2010/0326828 on theGenScript eStain 2.0 Protein Staining System using a blotting pad as agel-contacting stain solution carrier and destaining sink contact sheet(FIG. 3A), compared to using polyester cellulose material (FIG. 3B).

FIGS. 4A, B, C are pictures of electrophoresis gels processed using theprocedure of U.S. Patent Application Publication No. 2010/0326828 on theG2 Fast Blotting instrument using a blotting pad as a gel-contactingstain solution carrier and destaining sink contact sheet (FIG. 4A)compared to using polyester cellulose material (FIG. 4B, 4C).

FIGS. 5A, B, C, D are pictures of electrophoresis gels stained usingcommercially available Coomassie stain according to conventional methods(FIGS. 5C, 5D), compared to gels electrostained using polyestercellulose as the staining and destaining pads (FIGS. 5A, 5B) performedon the G2 Fast Blotting instrument.

A biopolymer includes, but is not limited to, a peptide, protein, RNA,DNA, single-stranded oligonucleotide, double-stranded oligonucleotide,oligosaccharide and any complexes, modifications, derivatives, etc.thereof. Examples of complexes include, but are not limited to,polypeptide-polypeptide complexes, RNA-polypeptide complexes,DNA-polypeptide complexes, polynucleotide-polynucleotide complexes, etc.

A matrix material includes any material in which a biopolymer isassociated with, impregnated within, bound to, absorbed to, the matrix.In one embodiment, the matrix is configured for use in an electricfield, typically a gel, e.g., SDS-polyacrylamide gel or native gel, butalso including a membrane, filter, etc. used in known electrophoresisprocesses. In one embodiment, the biopolymer is contained within cellsand/or tissue that are present on or associated with the matrix, such asa porous substrate, e.g., porous filter paper. In embodiments, the cellsare fixed using methods known in the art, and the tissue is tissuesections that may be fixed and prepared according to methods known inthe art.

The term sorbed includes adsorption, absorption, and the retention offluids between fibers, or within the porosity of or in hollow fibers,due to surface tension and related forces.

In one embodiment, the composition for staining or otherwise taggingbiopolymers in the matrix comprises a staining reagent, an acid, anorganic solvent, a staining agent, and at least one generally planarcontact sheet. In embodiments, the composition for staining may containadditives such as salts, chelating agents e.g., ethylene diaminetetraacetic acid (EDTA), fixatives, etc. In contrast to the porous,typically paper materials known in the art, the disclosed contact sheetis a fluid-permeable sheet primarily of a non-cellulosic material whichexhibits a substantially uniform, e.g., non-porous, surface in contactwith the matrix. In one embodiment, the sheet may consist primarily ofpolyester, and may be in the form of woven polyester fiber, or non-wovenpolyester fiber. In one embodiment, the sheet may consist primarily ofpolyester, but may also comprise a polyester cellulose blend. Forexample, the sheet or sheets may be in the form of a woven or non-wovenpolyester fiber and cotton cellulose fiber blend and/or felt. Inembodiments, the sheet or sheets may be primarily polyacrylamide,polysaccharide, and/or agarose. In embodiments, the sheet or sheets maybe primarily as described but include a mixture or blend with cellulose,alumina, silica, etc.

Specific examples of materials that can be used in the disclosedcomposition include woven polyester as in Contec Quiltec I wipes,non-woven polyester as in Contec SAT wipes or LymTech Scientific C3SHwipes, non-woven polyester cellulose as in LymTech Scientific C30Poly/Cellulose wipes, polyester cellulose, non-woven polyester celluloseas in Contec Amplitude Ecotech wipes, non-woven polypropylene as inLymTech Scientific 7408 Compo wipe including thermally bonded innercellulose layer, polypropylene and polypropylene blends, such as amulti-layered sheet having layers of polypropylene and inner layers ofcellulose, and polyacrylamide such as Pierce Precise Tris-Glycine gels.

The acid may be an organic acid, e.g., acetic acid, citric acid,sulfosalicylic acid, phosphoric acid, hydrochloric acid, sulfuric acid,etc., and may be at a concentration in the range of about 0.1% v/v toabout 20% v/v. In general, the acid may be any acid or mixture of acidsknown for use in electrophoretic separations, and is used to fixbiopolymers within an electrophoresis gel during execution of thestaining method. In one embodiment, the liquid phase of the compositionincludes five percent (5%) acetic acid.

The organic solvent may be a water soluble solvent, e.g., ethanol,methanol, isopropanol, etc., and may be a concentration in the range ofabout 1% v/v to about 50% v/v. In general, the organic solvent may beany solvent or mixture of solvents suitable for use with theelectrophoretic separation target(s) and a selected stain. In oneembodiment, the liquid phase of the composition includes 25% v/vethanol. In one embodiment, the liquid phase of the composition includes25% v/v isopropanol.

Optionally, the composition may include a buffering agent, such assodium phosphate, Tris, MES, MOPS, etc., and may be at a concentrationin the range of about 0.1% to about 10%, and at a pH of about pH 2 toabout pH 11. In one embodiment, the liquid phase of the compositionincludes sodium phosphate buffer.

In one embodiment, the composition is a source composition including astaining reagent, i.e., a stain. In one embodiment, the stain is aprotein stain, e.g., known protein stains including Coomassie Blue R-250or Coomassie Blue G-250, at a concentration range of about 0.01% w/v toabout 5% w/v. Other hydrophobic or charged stains including chromogenic,fluorescent, chemiluminescent, or bioluminescent molecules may be used,including stains for detecting protein modifications such asphosphorylation, glycosylation, acetylation, nitrosylation, SUMOylation,epitope tags, silver stains, and metal-protein complex stains. Otherstaining reagents used to tag biopolymers for other means of detection,such as non-protein biopolymers, include antibodies, nucleic acids,Fabs, scFv antibodies, aptamers, dendrimers, quantum dots, SYBR Green,SYTO 61, TOTO-3, ethidium bromide, propidium iodide, Hoechst, DAPI, etc.may be used. In one embodiment, the liquid phase of the compositionincludes or will include as subsequently described 0.01% w/v ofCoomassie Blue R-250.

In one embodiment, the acid, organic solvent, and staining reagent aresorbed to the fluid-permeable contact sheet. The composition may bepackaged in a fluid-impermeable bag (e.g., a plastic food storage bagwith a secure seal), sachet, overwrap, etc. that is removed prior toapplication of the composition to a gel. In one embodiment, thecomposition may further include a reservoir layer contactable with anon-matrix contact side of the contact sheet. The reservoir layer maycomprise a sorbent material, such as cellulose, that may hold asubstantial fraction of the acid, staining reagent, organic solvent,optional buffer, etc. for fluid communication through the contact sheetto the matrix. In one embodiment, the staining solution or a componentmay be dried onto the contact sheet and rehydrated before use.

In one embodiment, the composition is a sink composition including asolution, such as a destaining solution. In one embodiment, the destainsolution comprises an organic solvent, an acid, water, and optionallycomprises a buffering agent. In one embodiment, the sink composition ispaper. In one embodiment, the sink composition is the same sheetmaterial as the staining sheet but contains different describedsolutions. In one embodiment, the sink composition may further contain asorbent, such as cellulose, although at a lesser fraction than the non-or partially-cellulosic material that is incorporated into the contactsheet. In one embodiment, the sink composition may further contain asorbent such as alumina and silica. In one embodiment, the sinkcomposition may further include the destain solution and/or sorbent as alayer in contact with a non-matrix contact side of the sink contactsheet. The sorbent may preferentially sorb the staining reagent. Forexample, where the staining reagent is Coomassie Blue R-250, CoomassieBlue G-250, or a similar charged molecule, the sorbent may include ionexchange cellulose that preferentially binds organic anions.

As is apparent from the first and second constructions, the compositionneed not omit all cellulosic materials, but instead provides aprincipally non- or partially-cellulosic material layer that isdeposited in direct contact with the surface of the matrix, e.g.,electrophoresis gel. By replacing the cellulosic materials withessentially a non- or partially-cellulosic sheet, increasedstaining/destaining speed was retained while the stain background andblotchiness of an electrophoretically destained gel was dramaticallydecreased.

One embodiment is a kit containing a first composition including anacid, an organic solvent, a staining reagent, and a first generallyplanar, fluid-permeable contact sheet(s) consisting primarily of anon-cellulosic material. Any one or more of the acid, organic solvent,and staining reagent may be sorbed to the fluid-permeable contact sheet,may be sorbed to a reservoir layer contactable with non-gel contact sideof the first contact sheet, or may be provided separately. In oneembodiment, the kit further comprises a second composition including anacid, an organic solvent, a sorbent, and a generally planar,fluid-permeable contact sheet consisting primarily of a non-cellulosicmaterial. The sorbent may be a non-primary component of the contactsheet, or may be a sorbent layer contacting a non-gel contact side ofthe contact sheet. The acid, organic solvent, and non-cellulosicmaterial of the respective compositions need not be the same. Thesorbent may preferentially sorb the particular staining reagent of thefirst composition of the kit.

One embodiment is a method of staining a matrix using an electric field,such as an electrophoretic matrix, such as a gel, containingbiopolymers. After electrophoresis, the matrix with the separatedbiopolymers can be stained using pre-assembled non-cellulosic sheetsadsorbed with a staining solution (first composition; cathode sheets)and destaining solution (second composition; anode sheets), or it can bestained with non-cellulosic sheets adsorbed with a staining solution(first composition; cathode sheets) and destaining solution (secondcomposition; anode sheets) pre-adsorbed by the user.

In one embodiment, after electrophoresis, the gel was removed from theelectrophoresis cassette and pre-assembled solution-bearing sheets wereremoved from their packing and positioned on the appropriate sides ofthe electrophoresis gel with previously separated biopolymers. A DCvoltage was applied across the sheets-gel-sheets assembly for sevenminutes using Pierce Fast G2 Blotter (25 Volts, 1.0-1.3 Amp (constant))or eStain 2.0 Protein Staining System.

In one embodiment, a user places the non-cellulosic sheets into separatecontainers and adds 13-15 ml staining solution (first composition) and13-15 ml destaining solution (second composition) ensuring that thesolutions are evenly adsorbed into the sheets. After electrophoresis,the gel and pre-made solution-bearing sheets are positioned on theappropriate sides of an electrophoresis gel with previously separatedbiopolymers. A DC voltage is applied across the sheets-gel-sheetsassembly for seven minutes using Pierce Fast G2 Blotter (ThermoScientific) (25 Volts, 1.0-1.3 Amp (constant)) or eStain 2.0 ProteinStaining System.

EXAMPLE 1

FIGS. 1A-D show a comparison of SDS-PAGE gels stainedelectrophoretically using filter paper and by conventional Coomassiestaining protocol. Cell lysates, purified proteins, and molecular weightladders were loaded and separated by electrophoresis on polyacrylamidegels (FIG. 1A: 4%-12% NuPage Bis Tris; FIGS. 1B, D: 4%-20% CriterionTris-HCl; FIG. 1C: 4%-20% Midi Tris-Glycine) according to the gelsuppliers' recommendations. After electrophoresis, the midi gels werecut in half and mini gel was left as is. FIGS. 1A and B gels werestained using eStain Protein Staining Pad R-250 (GenScript L02011) whichuses filter paper for seven minutes according to the manufacturer'sinstructions. FIGS. 1 C and D gels were stained with Imperial ProteinStain (Thermo Scientific product #24615) by first washing the gel threetimes ten minutes with ultra-pure water and staining for 60 minutes inthe stain. The gel was destained overnight in water. These resultsshowed the uneven and splotchy staining artifacts produced duringelectrophoretic staining using conventional filter paper compared to theeven staining produced by Coomassie staining.

EXAMPLE 2

FIGS. 2A, B show comparison of the resistance (FIG. 2A) and temperaturegenerated (FIG. 2B) during the electrophoretic staining procedure.Biomolecules were separated on polyacrylamide gels as in FIG. 1 and thenprepared for electrophoretic staining by placing the gels between astaining pad of either polyester cellulose sheets or conventional filterpaper. The anode solution was 30% ethanol, 5% acetic acid and thecathode solution was 30% ethanol, 5% acetic acid, 0.01% Coomassie R-250.A DC voltage applied across the sheets-gel-sheets assembly for sevenminutes using Pierce Fast G2 Blotter (25 Volts, 1.0-1.3 Amp (constant)).The resistance and temperature were measured and plotted. These resultsdemonstrated lower resistance and heat generation during electrostainingusing polyester/cellulose material compared to filter paper.

EXAMPLE 3

FIGS. 3A, B show improved staining and destaining of polyacrylamide gelswhen electrostaining was performed using polyester cellulose material inthe staining and destaining pads compared to conventional filter paper.Protein samples were separated by electrophoresis on 4%-20% PreciseTris-Glycine polyacrylamide gels. After electrophoresis, both gels wereelectrostained using eStain 2.0 Protein Staining System. Cellulosefilter paper (FIG. 3A) or polyester cellulose material (LymTech C30L; 7layers) (FIG. 3B) were soaked in cathode-staining solution (25% ethanol,5% acetic acid, 0.01% Coomassie R-250) and anode-destaining solution(25% ethanol, 5% acetic acid) and placed on either side of the unstainedgel. The assembled sandwiches were placed into eStain Protein StainingSystem and the gels were electrostained for seven minutes. Thephotograph represents results from the inventive method showing lessblotchy staining and more even staining across the gel. In comparison toFIGS. 1 and 3A, FIG. 3B shows a lighter stain background of theprocessed electrophoresis gel and lacking the significant blotchingartifacts that appear in FIGS. 1 A and B particularly in the lowerquadrant, and in FIG. 3A particularly in the lower center area. Therewas thus significant improvement in destaining efficiency without resortto the passive, diffusion-driven staining and destaining processestypically used to obtain complete removal of stain background just byusing supple material, e.g., material that easily conforms to the givensurface, instead of more rigid filter paper.

EXAMPLE 4

FIGS. 4A-C show improved staining and destaining of polyacrylamide gelswith electrostaining using polyester cellulose compared to filter paperusing Thermo Scientific G2 Fast Blotter instrument. Protein samples wereseparated by electrophoresis on 4%-20% Criterion HCl (FIGS. 4A, 4B)polyacrylamide gel and 4%-20% Midi Tris-Glycine polyacrylamide gel (FIG.4C). After electrophoresis, both midi gels were cut in half andelectrostained using Pierce Fast G2 Blotter instrument. Cellulose filterpaper (FIG. 4A) or polyester cellulose material (LymTech C30L; 7 layers)(FIG. 4B, 4C) were soaked in cathode staining solution (25% ethanol, 5%acetic acid, 0.01% Coomassie R-250) and anode destaining solution (25%ethanol, 5% acetic acid) and placed on either side of the unstained gel.The assembled sandwiches were electrostained using Pierce Fast G2Blotter for seven minutes at 1.3 Amps (constant) and 25 Volts (limit).

EXAMPLE 5

FIGS. 5A-D show the superior performance of electrophoretic stainingusing polyester cellulose compared to conventional Coomassie staining.PageRuler Unstained Protein Ladder and/or HeLa lysate was loaded onto4%-20% Midi Tris-Glycine gel (FIGS. 5A, 5C) and 4-20% Criterion HCl gel(FIGS. 5B, 5D) together with bovine serum albumin (BSA) that wasserially diluted and prepared for SDS-PAGE. The gels were subjected toelectrophoresis according to the gel suppliers' recommendations. Afterelectrophoresis, the midi-sized gels were cut in half. Gel A waselectrostained using Pierce Fast G2 Blotter where a polyester cellulosematerial (LymTech C30L; 7 layers) soaked with an anode solution (30%isopropanol, 5% acetic acid) was placed on the anode plate, followed bythe unstained gel and a polyester cellulose material (LymTech C30L; 7layers) soaked with a cathode solution (30% isopropanol, 5% acetic acid,0.01% Coomassie R-250). Gel B was electrostained using Pierce Fast G2Blotter using a polyester cellulose material (LymTech C30L; 7 layers)soaked with an anode solution (35% ethanol, 5% acetic acid) was placedon the anode plate, followed by the unstained gel and a polyestercellulose material (LymTech C30L; 7 layers) soaked with a cathodesolution (35% ethanol, 5% acetic acid, 0.01% Coomassie R-250). Thecathode plate was placed on the top of the stacks and the cassettes wereinserted into the Pierce G2 Fast Blotter Control Unit for seven minuteswith constant amperage (1.3 Amps) and limited voltage (25 Volts). FIG.5C and FIG. 5D gels were stained with Imperial Protein Stain by firstwashing the gels three times fifteen minutes with ultra-pure water andstaining for sixty minutes in the stain. The gels were destainedovernight in water.

For comparison with conventional staining, the gel was conventionallystained using commercially available pre-formulated Coomassie stain,e.g., Pierce Imperial Protein Stain (FIGS. 1 and 5). Afterelectrophoresis the gel was removed from the cassette, washed threetimes for 10-15 minutes each with gentle agitation with ultra-purewater, and then stained for sixty minutes with gentle agitation. Thestaining solution was decanted and the gel was washed three times fortwenty minutes each in water or overnight with gentle agitation.

The embodiments shown and described in the specification are onlyspecific embodiments of inventors who are skilled in the art and are notlimiting in any way. Therefore, various changes, modifications, oralterations to those embodiments may be made without departing from thespirit of the invention in the scope of the following claims. Thereferences cited are expressly incorporated by reference herein in theirentirety.

What is claimed is:
 1. A method for electrophoretically stainingbiopolymers embedded in a matrix, the method comprising contacting afirst side of the matrix containing the biopolymer with a non-cellulosicfirst matrix contact sheet or a first matrix contact sheet including ablend containing cellulosic material, the first matrix contact sheethaving a matrix contact surface and a stain reagent, contacting a secondside of the matrix containing the biopolymer with a second matrixcontact sheet comprising a non- or partially-cellulosic material, havinga matrix contact surface and comprising a destain solution comprising anorganic solvent, an acid, water, and optionally a buffering agent, thecontacting of the first matrix contact sheet and the second matrixcontact sheet with the matrix forming a multi-layered assembly having athickness, and applying an electrical force across the thickness of theassembly sufficient to move the stain reagent into the matrix, therebystaining the biopolymer in the matrix, and to move excess free stainreagent out of the matrix, thereby destaining the matrix.
 2. The methodof claim 1 where the first matrix contact sheet further comprises atleast one additional sheet distal from the first side of the matrixand/or the second matrix contact sheet further comprises at least oneadditional sheet distal from the second side of the matrix, the at leastone additional sheet being the same or different from the first matrixcontact sheet and/or the second matrix contact sheet.
 3. The method ofclaim 1 where the first matrix contact sheet is capable of absorbing atleast 5 ml of a staining solution, but less than 15 ml of the stainingsolution; a thickness of the first matrix contact sheet is less than 0.5inches; and/or the matrix contact surface is sufficient to providemaximum contact between the first matrix contact sheet and the matrix.4. The method of claim 1 where the first matrix contact sheet or thefirst and the second matrix contact sheet is polyester,polyester/cellulose blend, polypropylene, polypropylene/cellulose blend,polyacrylamide, nitrocellulose, non-cellulose, polysaccharide, agarose,felt, polyvinylidene fluoride (PVDF), polyvinylidene difluoride, finewoven cotton, wool, regenerated cellulose fiber, Lyocell, andcombinations thereof.
 5. The method of claim 4 where the first matrixcontact sheet is polyester, polyester/cellulose blend, polypropylene,polypropylene/cellulose blend, polyacrylamide, nitrocellulose,non-cellulose, polysaccharide, agarose, felt, polyvinylidene fluoride(PVDF), polyvinylidene difluoride, regenerated cellulose fiber, Lyocell,and combinations thereof.
 6. The method of claim 2 where the at leastone additional sheet is polyester, polyester/cellulose blend,polyacrylamide, nitrocellulose, polysaccharide, agarose, felt,polyvinylidene fluoride (PVDF), polyvinylidene difluoride, a fabric, afoam, a sponge, and combinations thereof.
 7. The method of claim 2 wherea stain solution and/or the destain solution is present in the at leastone additional sheet which is not in contact with the matrix.
 8. Themethod of claim 1 where the second matrix contact sheet comprises asorbent selected from the group consisting of wound dressingnon-adhesive material, woven and non-woven cotton, foam, felt, agarose,agarose/polyacrylamide matrix, alumina, silica, polysaccharides,polymer, and combinations thereof.
 9. The method of claim 1 furthercomprising, prior to contacting the matrix with the first and secondmatrix contacting sheet, contacting the first matrix contacting sheetwith a stain solution, such that the stain solution is absorbed into thefirst matrix contact sheet, and contacting the second matrix contactingsheet with the destain solution, such that the destain solution isabsorbed into the second matrix contact sheet.
 10. The method of claim 1further comprising a staining solution, where the staining solutioncomprises an acid, an organic solvent, and the staining reagent.
 11. Themethod of claim 10 where the acid is acetic acid and the organic solventis ethanol or isopropanol.
 12. The method of claim 10 where the stainingsolution further comprises a buffering agent.
 13. The method of claim 12where the buffering agent is sodium phosphate.
 14. The method of claim 1where the staining reagent is a protein stain, and is selected from thegroup consisting of Coomassie Blue R-250, Coomassie Blue G-250, silverstain, hydrophobic or charged staining reagents, neutral stainingreagents, and protein modification stains.
 15. The method of claim 14where the charged staining reagents include chromogenic, fluorescent,chemiluminescent, and/or bioluminescent molecules.
 16. The method ofclaim 14 where the neutral staining reagent further comprises anon-charged organic or non-organic solvent.
 17. The method of claim 14where the protein modification is at least one of phosphorylation,glycosylation, acetylation, nitrosylation, SUMOylation, or epitope tags.18. The method of claim 1 further comprising a stain solution, where thestain solution comprises 25% ethanol, 5% acetic acid, and 0.01%Coomassie Brilliant Blue dye R-250 or G-250, and the destain solutioncomprises 25% ethanol and 5% acetic acid.
 19. The method of claim 1where the matrix is selected from the group consisting ofSDS-polyacrylamide gels, native polyacrylamide gels, membranes, andfilters.
 20. A method for electrophoretically staining biopolymersembedded in a matrix, the method comprising forming an electrophoretic,multi-layered assembly by contacting a first side of the matrixcontaining the biopolymer with a first matrix contact sheet consistingprimarily of a non-cellulosic or partially cellulosic material having amatrix contact side in direct contact with the first side of the matrix,the first matrix contact sheet comprising a stain solution comprising astain reagent and a solvent, contacting a second side of the matrixcontaining the biopolymer with a second matrix contact sheet comprisinga non- or partially-cellulosic material having a matrix contact side indirect contact with the second side of the matrix, the second matrixcontact sheet comprising a destain solution comprising an organicsolvent, an acid, water, and optionally a buffering agent, positioningthe assembly between a first electrode and a second electrode and inelectrical contact with said electrodes; and applying a current to theelectrodes and across the assembly to provide an electric forcesufficient to move the staining reagent into the matrix, therebystaining the biopolymer in the matrix, and to move excess free stainingreagent out of the matrix, thereby destaining the matrix.
 21. The methodof claim 20 where the first electrode is a cathode and is stainlesssteel or titanium and the second electrode is an anode and isplatinum-coated titanium or carbon.